Ebola virus (EBOV) is an enveloped, nonsegmented, negative-strand RNA virus within the filovirus family, which can cause a lethal hemorrhagic febrile disease in humans and nonhuman primates, killing up to 90% of those, infected. There are currently no effective antiviral agents available for the treatment of EBOV infections nor have appropriate vaccines been developed for human disease prevention. The events of September 11, 2001 and the ensuing inhalation anthrax bioterrorism incidents have highlighted concerns that EBOV also might be used as a bioterrorism agent that could be disseminated via the aerosol route. However, little is known about the capacity of EBOV to colonize the human lower respiratory tract and to subsequently induce a systemic viral hemorrhagic fever syndrome. There are currently no published in vitro models for studying the effects of EBOV on human lung cells. Our preliminary data indicate that EBOV can replicate in NORMAL HUMAN BRONCHIAL EPITHELIAL CELLS (HBEC) maintained in air/liquid interface primary culture. In this R21 application, we propose that: inhaled EBOV aerosols are able to colonize the lungs by establishing a persistent infection in bronchial epithelium, leading to subsequent infection of alveolar macrophages. The SPECIFIC HYPOTHESIS is that EBOV entry into the bronchial epithelium is associated with plasma membrane-associated caveolae in these cells. Furthermore, once entry has occurred, viral replication within the bronchial epithelium (which requires the EBOV RNA-dependent RNA polymerase L and glycoprotein GP genes) induces up-regulation of interleukin-1 (IL-1), cyclooxygenase-2 (COX-2), and the inducible form of nitric oxide synthase (iNOS), which, in turn, facilitate further viral replication and inhibit bronchial epithelial apoptosis. To test this hypothesis both in vitro and in vivo, we propose the following four SPECIFIC AIMS: (1) to determine whether HBEC are permissive for EBOV entry and replication without inducing apoptosis and/or cytotoxicity; (2) to determine whether caveolae are required for cellular entry into HBEC; (3) to determine whether IL-1-mediated up-regulation of COX-2 and iNOS expression in HBEC prevent apoptosis and facilitate EBOV replication within HBEC; (4) to determine whether RNA interference (RNAi), by inducing post-transcriptional silencing of EBOV GP and L genes, can prevent or inhibit the replication and budding of virus within HBEC. These studies are expected to provide new information as to how the entry, replication, and budding of EBOV in HBEC could be modulated by multiple targeted biodefense approaches against aerosolized EBOV.